Genomewide association analysis of coronary artery disease

Background - Modern genotyping platforms permit a systematic search for inherited components of complex diseases. We performed a joint analysis of two genomewide association studies of coronary artery disease. Methods - We first identified chromosomal loci that were strongly associated with coronary artery disease in the Wellcome Trust Case Control Consortium (WTCCC) study (which involved 1926 case subjects with coronary artery disease and 2938 controls) and looked for replication in the German MI [Myocardial Infarction] Family Study (which involved 875 case subjects with myocardial infarction and 1644 controls). Data on other single-nucleotide polymorphisms (SNPs) that were significantly associated with coronary artery disease in either study (P<0.001) were then combined to identify additional loci with a high probability of true association. Genotyping in both studies was performed with the use of the GeneChip Human Mapping 500K Array Set (Affymetrix). Results - Of thousands of chromosomal loci studied, the same locus had the strongest association with coronary artery disease in both the WTCCC and the German studies: chromosome 9p21.3 (SNP, rs1333049) (P=1.80x10–14 and P=3.40x10–6, respectively). Overall, the WTCCC study revealed nine loci that were strongly associated with coronary artery disease (P80%) of a true association: chromosomes 1p13.3 (rs599839), 1q41 (rs17465637), 10q11.21 (rs501120), and 15q22.33 (rs17228212). Conclusions - We identified several genetic loci that, individually and in aggregate, substantially affect the risk of development of coronary artery disease

Integrative pathway genomics of lung function and airflow obstruction

Chronic respiratory disorders are important contributors to the global burden of disease. Genome-wide association studies (GWASs) of lung function measures have identified several trait-associated loci, but explain only a modest portion of the phenotypic variability. We postulated that integrating pathway-based methods with GWASs of pulmonary function and airflow obstruction would identify a broader repertoire of genes and processes influencing these traits. We performed two independent GWASs of lung function and applied gene set enrichment analysis to one of the studies and validated the results using the second GWAS. We identified 131 significantly enriched gene sets associated with lung function and clustered them into larger biological modules involved in diverse processes including development, immunity, cell signalling, proliferation and arachidonic acid. We found that enrichment of gene sets was not driven by GWAS-significant variants or loci, but instead by those with less stringent association P-values. Next, we applied pathway enrichment analysis to a meta-analysed GWAS of airflow obstruction. We identified several biologic modules that functionally overlapped with those associated with pulmonary function. However, differences were also noted, including enrichment of extracellular matrix (ECM) processes specifically in the airflow obstruction study. Network analysis of the ECM module implicated a candidate gene, matrix metalloproteinase 10 (MMP10), as a putative disease target. We used a knockout mouse model to functionally validate MMP10’s role in influencing lung’s susceptibility to cigarette smoke-induced emphysema. By integrating pathway analysis with population-based genomics, we unravelled biologic processes underlying pulmonary function traits and identified a candidate gene for obstructive lung disease

Genomic copy number variation, human health, and disease

Despite the long recognised effects of chromosomal structural abnormalities and completion of the Human Genome Project, much of the structural variation in the genome has gone unrecognised until recently. Deletions and duplications of DNA strands of between a few hundred bp and several million bp—collectively referred to as copy number variants—are now known to be widespread. Since 2007, rigorous and adequately powered genome-wide association studies based on single nucleotide polymorphisms have yielded replicated associations to several common diseases. Some copy number variants explain rare, previously uncharacterised disorders, and they are now expected to explain some of the genetic contribution to common diseases. We review efforts to map copy number variants and discuss present and future prospects for assessment of their relation to human health and disease

Understanding the impact of pre-analytic variation in haematological and clinical chemistry analytes on the power of association studies

BACKGROUND: Errors, introduced through poor assessment of physical measurement or because of inconsistent or inappropriate standard operating procedures for collecting, processing, storing or analysing haematological and biochemistry analytes, have a negative impact on the power of association studies using the collected data. A dataset from UK Biobank was used to evaluate the impact of pre-analytical variability on the power of association studies. METHODS: First, we estimated the proportion of the variance in analyte concentration that may be attributed to delay in processing using variance component analysis. Then, we captured the proportion of heterogeneity between subjects that is due to variability in the rate of degradation of analytes, by fitting a mixed model. Finally, we evaluated the impact of delay in processing on the power of a nested case-control study using a power calculator that we developed and which takes into account uncertainty in outcome and explanatory variables measurements. RESULTS: The results showed that (i) the majority of the analytes investigated in our analysis, were stable over a period of 36 h and (ii) some analytes were unstable and the resulting pre-analytical variation substantially decreased the power of the study, under the settings we investigated. CONCLUSIONS: It is important to specify a limited delay in processing for analytes that are very sensitive to delayed assay. If the rate of degradation of an analyte varies between individuals, any delay introduces a bias which increases with increasing delay. If pre-analytical variation occurring due to delays in sample processing is ignored, it affects adversely the power of the studies that use the data

Tumour necrosis factor gene complex polymorphisms and haplotypes in chronic obstructive pulmonary disease

We aimed to examine the role of tumour necrosis factor gene complex polymorphisms in subjects with chronic obstructive pulmonary disease (COPD). We hypothesized that individuals possessing polymorphic variants associated with higher TNF secretion would be more susceptible to and/or have more severe disease. Patients with COPD and population controls underwent detailed clinical phenotyping. Genotyping for the tumour necrosis factor-308 and the lymphotoxin alpha NcoI (LTα) polymorphisms was carried out by ‘blinded’ laboratory staff. 361 individuals (220 cases and 141 controls) were recruited. We showed an association between the LTαNcol polymorphism and forced vital capacity (FVC) in a population of older adults with and without COPD. The LTαNcol*2 allele was associated with poorer lung function, under a codominant model, with a fall in FVC (expressed as a percentage of its predicted value) of 3.7% for each copy of the LTαNcol*2 allele possessed (for FVC, regression coefficient (95% CI) = -3.73 (-7.01to -0.44), p=0.026; for FEV1 regression coefficient = -3.56 (-7.80 to 0.70), p = 0.101. However, there was no difference in genotype distribution between the case and control populations. This study adds weight to the suggestion that the TNF gene complex is involved in physiological alterations (FVC) that may affect the development and severity of COPD. The absence of a significant association between the TNF gene-complex polymorphisms in this study does not rule out a modest effect of these polymorphisms on the risk of COPD, as much larger studies are needed to detect modest gene effects on binary disease endpoints

Human exome-chip meta-analysis identifies novel genetic loci associated with smoking behaviour

Smoking is a major risk factor for many diseases, including common respiratory disorders such as chronic obstructive pulmonary disease. Previous GWASs have been successful in identifying 13 common SNPs associated with smoking behaviour. Meta-analysing summary data from up to 33 cohorts, we have carried out an association study between the SNPs found on the exome-chip (n ≈250k SNPs), which predominantly assays rare putatively functional variants, and four traits: Smoking Initiation (n≈80k), Smoking Cessation (n≈41.5k), Cigarettes Per Day (n≈26.5k) and Pack Years (n≈33k). In addition to identifying previously reported signals with regards to smoking behaviour, we have identified SNPs in 5 novel regions, including a rare variant on chromosome 16 (MAF=0.01%). The previously identified SNPs fall within CHRNA5 (rs16969968, missense), CHRNA3 (rs938682, intronic) and IREB2 (rs13180, synonymous) – which are all located at the 15q25 locus. We are currently initiating follow-up studies; and if replicated, the novel loci identified in this study will facilitate understanding the genetic aetiology of smoking behaviour and may lead to identification of drug targets of potential relevance to smoking cessation

Human CCL3L1 copy number variation, gene expression, and the role of the CCL3L1-CCR5 axis in lung function

Background: The CCL3L1-CCR5 signaling axis is important in a number of inflammatory responses, including macrophage function, and T-cell-dependent immune responses. Small molecule CCR5 antagonists exist, including the approved antiretroviral drug maraviroc, and therapeutic monoclonal antibodies are in development. Repositioning of drugs and targets into new disease areas can accelerate the availability of new therapies and substantially reduce costs. As it has been shown that drug targets with genetic evidence supporting their involvement in the disease are more likely to be successful in clinical development, using genetic association studies to identify new target repurposing opportunities could be fruitful. Here we investigate the potential of perturbation of the CCL3L1-CCR5 axis as treatment for respiratory disease. Europeans typically carry between 0 and 5 copies of CCL3L1 and this multi-allelic variation is not detected by widely used genome-wide single nucleotide polymorphism studies.  Methods: We directly measured the complex structural variation of CCL3L1 using the Paralogue Ratio Test and imputed (with validation) CCR5del32 genotypes in 5,000 individuals from UK Biobank, selected from the extremes of the lung function distribution, and analysed DNA and RNAseq data for CCL3L1 from the 1000 Genomes Project. Results: We confirmed the gene dosage effect of CCL3L1 copy number on CCL3L1 mRNA expression levels.  We found no evidence for association of CCL3L1 copy number or CCR5del32 genotype with lung function. Conclusions: These results suggest that repositioning CCR5 antagonists is unlikely to be successful for the treatment of airflow obstruction

Genetic analysis of over one million people identifies 535 novel loci for blood pressure

High blood pressure is a highly heritable and modifiable risk factor for cardiovascular disease. We report the largest genetic association study of blood pressure traits (systolic, diastolic, pulse pressure) to date in over one million people of European ancestry. We identify 535 novel blood pressure loci that not only offer new biological insights into blood pressure regulation but also reveal shared genetic architecture between blood pressure and lifestyle exposures. Our findings identify new biological pathways for blood pressure regulation with potential for improved cardiovascular disease prevention in the future

Slamming the door on trade policy discretion? : the WTO Appellate Body’s ruling on market distortions and production costs in EU-Biodiesel (Argentina)

This paper presents a legal-economic analysis of the Appellate Body’s decision that the WTO’s Anti-Dumping Agreement (ADA) precludes countries from taking into account government-created price distortions of major inputs when calculating anti-dumping duties, made in EU-Biodiesel (Argentina). In this case, the EU made adjustments to the price of biodiesel’s principal input – soybeans – in determining the cost of production of biodiesel in Argentina. The adjustment was made based on the uncontested finding that the price of soybeans in Argentina was distorted by the existence of an export tax scheme that resulted in artificially low soybean prices. The Appellate Body found that the EU was not permitted to take tax policy-induced price distortions into account in calculating dumping margins. We analyze the economic rationale for Argentina’s export tax system, distortions in biodiesel markets in Argentina and the EU, and the remaining trade policy options for addressing distorted international prices. We also assess whether existing subsidies disciplines would be more effective in addressing this problem and conclude that they would not

High-Throughput Sequencing in Respiratory, Critical Care, and Sleep Medicine Research An Official American Thoracic Society Workshop Report

High-throughput, “next-generation” sequencing methods are now being broadly applied across all fields of biomedical research, including respiratory disease, critical care, and sleep medicine. Although there are numerous review articles and best practice guidelines related to sequencing methods and data analysis, there are fewer resources summarizing issues related to study design and interpretation, especially as applied to common, complex, nonmalignant diseases. To address these gaps, a single-day workshop was held at the American Thoracic Society meeting in May 2017, led by the American Thoracic Society Section on Genetics and Genomics. The aim of this workshop was to review the design, analysis, interpretation, and functional follow-up of high-throughput sequencing studies in respiratory, critical care, and sleep medicine research. This workshop brought together experts in multiple fields, including genetic epidemiology, biobanking bioinformatics, and research ethics, along with physician-scientists with expertise in a range of relevant diseases. The workshop focused on application of DNA and RNA sequencing research in common chronic diseases and did not cover sequencing studies in lung cancer, monogenic diseases (e.g., cystic fibrosis), or microbiome sequencing. Participants reviewed and discussed study design, data analysis and presentation, interpretation, functional follow-up, and reporting of results. This report summarizes the main conclusions of the workshop, specifically addressing the application of these methods in respiratory, critical care, and sleep medicine research. This workshop report may serve as a resource for our research community as well as for journal editors and reviewers of sequencing-based manuscript submissions in our research field